Field of the Invention
This invention relates to a test piece for use in DNA analysis, immunological analysis, and the like, and a system for reading out image information from the test piece.
Description of the Related Art
Recently, genetic engineering has exhibited rapid progress, and the human genome project for decoding the base sequence of human genomes which amount to 100,000 in number is progressing.
Further, enzyme immunoassay, fluorescent antibody technique and the like utilizing antigen-antibody reactions have been used in diagnoses and studies, and studies for searching DNAs which affect genetic diseases are now progressing. In such a situation, a microarray technique is now attracting attention.
In the microarray technique, a microarray chip (sometimes called a DNA chip) comprising a plurality of known cDNAs (an example of specific binding substances) coated in a matrix on a substrate such as a membrane filter or a slide glass at a high density (at intervals of not larger than several hundred xcexcm) is used and DNAs (an example of organism-originating substances) taken from cells of a normal person A and labeled with a fluorescent dye a and DNAs taken from cells of a genetic-diseased person B and labeled with a fluorescent dye b are dropped onto the microarray chip by pipettes or the like, thereby hybridizing the DNAs of the specimens with the cDNAs on the microarray chip. Thereafter, exciting light beams which respectively excite the fluorescent dyes a and b are projected onto the cDNAs by causing the exciting light beams to scan the microarray chip and fluorescence emitted from each other of the cDNAs is detected by a photodetector. Then the cDNAs with which the DNAs of each specimen are hybridized are determined on the basis of the result of the detection, and the cDNAs with which the DNAs of the normal person A are hybridized and those with which the DNAs of the diseased person B are hybridized are compared, whereby DNAs expressed or lost by genetic disease can be determined.
In the microarray technique, it is necessary to precisely two-dimensionally scan the microarray chip coated with cDNAs at a high density, and there has been proposed a radiation image read-out apparatus with such a precise scanning system. See, for instance, Japanese Unexamined Patent Publication No. 10(1998)-3134.
The kinds of cDNAs to be used sometimes amount to several tens of thousands and in such a case, the cDNAs must be coated on a plurality of substrates. However when the number of the microarray chips to be used increases, replacement of microarray chips becomes troublesome.
In view of the foregoing observations and description, the primary object of the present invention is to provide a test piece on which an increased number of specific binding substances such as cDNAs can be disposed, and a system for reading out image information from the test piece.
In accordance with one aspect of the present invention, there is provided a test piece such as a microarray chip for use in biological analyses comprising a plurality of different known specific binding substances such as cDNAs disposed in predetermined positions on a substrate such as a slide glass, wherein the improvement comprises that
the specific binding substances are disposed on a plurality of surfaces provided by the substrate and arranged in the direction of thickness of the substrate.
The plurality of surfaces provided by the substrate and arranged in the direction of thickness of the substrate may be opposite sides of the substrate or may be provided by a multi-layered substrate formed by a plurality of substrates which are stacked and bonded together so that the surfaces on which the specific binding substances are disposed are substantially in parallel to each other.
It is preferred that the specific binding substances be disposed on the surfaces in positions where the specific binding substances on the respective surfaces do not interfere with each other in the direction of thickness of the substrate, that is, the specific binding substances on the respective surfaces do not overlap with each other in the direction of thickness of the substrate.
The substrate may be formed of any material so long as the specific binding substances can be spotted and stably held on the substrate and the substrate is optically transparent to the exciting light and the fluorescence emitted from the specific binding substances upon exposure to the exciting light. For example, the substrate may be a membrane filter or a slide glass. Further the substrate may be subjected to pretreatment so that the specific binding substances are stably held on the substrate.
The specific binding substances include hormones, tumor markers, enzymes, antibodies, antigens, abzymes, other proteins, nucleic acids, cDNAs, DNAs, RNAs, and the like, and means those which can be specifically bound with an organism-originating substance. The means of the expression xe2x80x9cknownxe2x80x9d differs by the specific binding substance. For example, when the specific binding substance is a nucleic acid, xe2x80x9cknownxe2x80x9d means that the base sequence, the lengths of the bases and the like are known, and when specific binding substance is protein, xe2x80x9cknown xe2x80x9d means that the composition of the amino acid is known. The specific binding substances are disposed by one kind for each position.
In accordance with another aspect of the present invention, there is provided a system for reading out image information from the test piece of the present invention comprising
a test piece holder portion which holds a test piece of the present invention the specific binding substances on which have been hybridized with an organism-originating substance labeled with fluorescent dye,
an exciting light source which emits exciting light for exciting the fluorescent dye,
a photoelectric read-out means which photoelectrically reads out fluorescence emitted from the fluorescent dye upon exposure to the exciting light,
a scanning means which has an optical head for projecting the exciting light onto the test piece and leading fluorescence, which is emitted from the fluorescent dye and travels through the surface of the test piece onto which the exciting light is projected, to the photoelectric read-out means, and causes the exciting light to scan the test piece, and
a controller which controls the exciting light source, the photoelectric read-out means and the scanning means so that fluorescence emitted from the specific binding substances upon exposure to the exciting light is detected for each of the surfaces of the test piece.
The test piece holder portion may comprise a table on which the test piece is placed. In this case, the test piece is placed on the table with its one side in contact with the table, and accordingly, it is necessary that the table is transparent to at least the fluorescence. When the test piece holder portion is in the form of a member which supports only the four corners of the test piece, the test piece holder portion need not be transparent.
The organism-originating substance may be a wide variety of substances originated from an organism including hormones, tumor markers, enzymes, proteins, antibodies, various substances which can be antigens, nucleic acids, cDNAs, mRNAs and the like.
The exciting light is light suitable for exciting the fluorescent dye including a laser beam.
As the photoelectric read-out means, a photomultiplier which can detect at a high sensitivity weak light such as fluorescence may be suitably used. However, various known photoelectric read-out means such as a cooled CCD may be used without limited to the photomultiplier.
In accordance with the present invention, since the specific binding substances are disposed on a plurality of surfaces provided by the substrate and arranged in the direction of thickness of the substrate, an increased number of specific binding substances can be disposed on one test piece and accordingly, the number of test pieces to be used can be less even if a large number of specific binding substances are used, whereby the frequency at which the test pieces are replaced can be reduced and reading operation can be effectively performed.
When the specific binding substances are disposed on opposite sides of a single substrate, the test piece can be manufactured at low cost.